Fiber‐optic spectrophotometry of cowpea nodules treated with nitrate or ammonium

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Abstract
Cowpea nodules (Vigna unguiculata [L.] Walp. cv. CB5) were pierced with two opposing optical fibers (280 μm diameter), and absorbance spectra of 0.1 mm of nodule tissue were recorded from 415 to 600 nm using a modified spectrophotometer with a nodule sampling stage. The nodule spectra exhibited two absorbance bands, a major band in the near‐UV (415–450 nm) and a lesser one in the green‐yellow region (510–585 nm); the latter exhibited a prominent peak at 550 nm. Nodule spectra were consistent with the superposition of the spectra of ferroleghemoglobin (Lb2+), oxyleghemoglobin (Lb2+‐ O2), and cytochrome c (550 nm). The detection of leghemoglobin in vivo was confirmed by demonstrating the reversibility of binding of CO to Lb, and by comparing the spectra of live nodules with those obtained from anaerobic leghemoglobin preparations. The effects on the nodule spectra of two successive applications (36 and 39 days after planting) of 5 and 10 mM NO3 ‐ or NH4 + to the nutrient solution bathing the nodulated roots were determined. The spectra of NO3 ‐‐ and NH4 +‐treated nodules were indistinguishable; in each case the height of both the near‐UV and green‐yellow absorbance bands decreased with treatment. Treatment with N caused a significant reduction in the area of the green‐yellow absorbance band. The spectra of green leghemoglobin pigments were obtained from senescent nodules; these exhibited a reduction in the height of the near‐UV absorbance band and a flattening of the green‐yellow band similar to that of nodules treated with N, but the broadening of the near‐UV band was greater in green nodules.